Biomedical subjects
Publications and source records attributed to A van Belkum.
The main mechanisms by which Campylobacter jejuni causes diarrhea are unknown. In contrast to a recent communication, we report here the absence of an association with the plasmid pVir in patients infected with C. jejuni who developed bloody diarrhea in The Netherlands, and we suggest a role for other virulence determinants.
We investigated the prevalence and determinants of nasopharyngeal carriage of Neisseria meningitidis in 3200 healthy children aged 1-19 years. The incidence of meningococcal carriage was, on average, 1.5%. Peak incidences were seen at age 1 year and after age 15 years. The independent determinants of meningococcal carriage included age, regular visits to youth clubs (odds ratio [OR], 2.2) and discotheques (OR, 4.3), and pneumococcal carriage (OR, 4.1).
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The performances of the BDProbeTec ET (Becton Dickinson) and COBAS AMPLICOR MTB (Roche) were retrospectively evaluated for detecting Mycobacterium tuberculosis complex in various respiratory specimens. The BACTEC and MGIT liquid culture system (Becton Dickinson) was used as a reference method. A total of 824 respiratory specimens, comprised of sputa, bronchoalveolar lavage fluid, and bronchial and tracheal aspirates from 580 patients, were evaluated. Out of 824 clinical specimens, 109 specimens from 43 patients were culture positive for M. tuberculosis. Of these 109 specimens, 67 were smear positive, 85 were positive by the COBAS AMPLICOR MTB test, and 94 were positive by the BDProbeTec ET. Of the 715 culture-negative specimens, 17 were positive by the auramine staining, 11 were positive by the COBAS AMPLICOR MTB test, and 12 were positive by the BDProbeTec ET. After discrepancy analysis and review of the patients' clinical data, 130 specimens from 50 patients were considered "true-positive" specimens. This resulted in the following sensitivities: microscopy, 61.5%; COBAS AMPLICOR MTB test, 78.0%; and BDProbeTec ET, 86.2%. The specificities of each system, based on the clinical diagnosis, were 99.7% for microscopy, 99.9% for the COBAS AMPLICOR MTB test, and 99.9% for the BDProbeTec ET. The data presented represent a considerable number of specimens evaluated with a considerable number of culture- and auramine-positive and culture-positive and auramine-negative results and therefore give a realistic view of how the data should be interpreted in a daily routine situation. Specifically, the data with regard to the culture-positive and auramine-negative specimens are useful, because in a routine situation, auramine-negative specimens are sometimes accepted, on clinical indications, to be analyzed by an amplification method.
The success of large-scale screening for Chlamydia trachomatis depends on the availability of noninvasive samples, low costs, and high-quality testing. To evaluate C. trachomatis testing with pregnant women, first-void urine specimens from 750 consecutive asymptomatic pregnant women from the Rotterdam area (The Netherlands) were collected. Initially, we investigated the performance of three different DNA isolation methods with 350 of these urines and 70 pools of 5 of the same subset of urine samples. The routinely used COBAS AMPLICOR test was compared to the COBAS AMPLICOR test with prior DNA isolation by use of the MagNA Pure large-volume kit and the MagNA Pure bacterial DNA isolation kit. The latter combination provided the best DNA test for pooled urines, with a sensitivity twice that of the other methods. Next, using all 750 urines, the COBAS AMPLICOR performance for individual testing was compared to pooled testing with the standard COBAS AMPLICOR procedure and subsequently to pooled testing with COBAS AMPLICOR in combination with the MagNA Pure bacterial DNA isolation kit. The sensitivity of COBAS AMPLICOR was 65% on individual and 42% on pooled urines but improved to 92% on pooled urines with the MagNA Pure bacterial DNA isolation kit, making this combination the best screening method. The C. trachomatis prevalence in this population appeared to be 6.4%. Additionally, the cost of the combined MagNA Pure bacterial DNA isolation kit and COBAS AMPLICOR method on pooled urines was only 56% of the cost of the standard COBAS AMPLICOR test applied to individual urines. Costs per positive case detected in the combined method were 39% of standard costs.
A trial with a 7-valent pneumococcal-conjugate vaccine in children with recurrent acute otitis media showed a shift in pneumococcal colonisation towards non-vaccine serotypes and an increase in Staphylococcus aureus-related acute otitis media after vaccination. We investigated prevalence and determinants of nasopharyngeal carriage of Streptococcus pneumoniae and S aureus in 3198 healthy children aged 1-19 years. Nasopharyngeal carriage of S pneumoniae was detected in 598 (19%) children, and was affected by age (peak incidence at 3 years) and day-care attendance (odds ratio [OR] 2.14, 95% CI 1.44-3.18). S aureus carriage was affected by age (peak incidence at 10 years) and male sex (OR 1.46, 1.25-1.70). Serotyping showed 42% vaccine type pneumococci. We noted a negative correlation for co-colonisation of S aureus and vaccine-type pneumococci (OR 0.68, 0.48-0.94), but not for S aureus and non-vaccine serotypes. These findings suggest a natural competition between colonisation with vaccine-type pneumococci and S aureus, which might explain the increase in S aureus-related otitis media after vaccination.
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Staphylococcus aureus is a common human pathogen in community- and hospital-acquired infections and its capsule is involved in pathogenesis. We report here the identification of type-5 and type-8 capsular antigens of S. aureus and the prevalence of such strains among volunteers in various age and population groups from different locations in India. S. aureus carriage rates varied between 18 and 50% with the highest values among university students and the lowest in schoolchildren, aged 6-20 years. There was no difference in carriage rates for males vs. females (P=0.415) or in the socioeconomic status of carriers vs. non-carriers or age dependence. Among the carriage isolates 21% were type-5, 52% were type-8 and the remaining 27% were non-typable. Among invasive isolates these percentages were 6, 64 and 30 respectively. This implies that type-5 strains may be less invasive than type-8 strains (P=0.0015).
The genus Madurella, described for non-sporulating agents of human mycetoma, is proven to be heterogeneous on the basis of rDNA small subunit (SSU) and Internal Transcribed Spacer (ITS) sequencing data. Madurella mycetomatis, the main agent of mycetoma in arid zones of Central and East Africa, probably belongs to the ascomycete order Sordariales. Madurella mycetomatis, the generic type species, is neotypified. Madurella grisea, with worldwide occurrence, is likely to be a member of the order Pleosporales, just as the mycetoma agents of Leptosphaeria, Pseudochaetosphaeronema, and Pyrenochaeta. Neotestudina rosatii belongs to the order Dothideales. Judging from ITS data, M. mycetomatis and N. rosatii are species complexes. The ex-type strain of N. rosatii, from a human mycetome, has an ITS sequence that deviates from that of environmental strains of the species.
Molecular methods for the rapid identification of methicillin-resistant Staphylococcus aureus (MRSA) are generally based on the detection of an S. aureus-specific gene target and the mecA gene. However, such methods cannot be applied for the direct detection of MRSA from nonsterile specimens such as nasal samples without the previous isolation, capture, or enrichment of MRSA because these samples often contain both coagulase-negative staphylococci (CoNS) and S. aureus, either of which can carry mecA. In this study, we describe a real-time multiplex PCR assay which allows the detection of MRSA directly from clinical specimens containing a mixture of staphylococci in <1 h. Five primers specific to the different staphylococcal cassette chromosome mec (SCCmec) right extremity sequences, including three new sequences, were used in combination with a primer and three molecular beacon probes specific to the S. aureus chromosomal orfX gene sequences located to the right of the SCCmec integration site. Of the 1,657 MRSA isolates tested, 1,636 (98.7%) were detected with the PCR assay, whereas 26 of 569 (4.6%) methicillin-susceptible S. aureus (MSSA) strains were misidentified as MRSA. None of the 62 nonstaphylococcal bacterial species or the 212 methicillin-resistant or 74 methicillin-susceptible CoNS strains (MRCoNS and MSCoNS, respectively) were detected by the assay. The amplification of MRSA was not inhibited in the presence of high copy numbers of MSSA, MRCoNS, or MSCoNS. The analytical sensitivity of the PCR assay, as evaluated with MRSA-negative nasal specimens containing a mixture of MSSA, MRCoNS, and MSCoNS spiked with MRSA, was approximately 25 CFU per nasal sample. This real-time PCR assay represents a rapid and powerful method which can be used for the detection of MRSA directly from specimens containing a mixture of staphylococci.
Mapping, and ultimately preventing, the dissemination of infectious agents is an important topic in public health. Newly developed molecular-microbiological methods have contributed significantly to recent advances in the efficient tracking of the nosocomial and environmental spread of microbial pathogens. Not only has the application of novel technologies led to improved understanding of microbial epidemiology, but the concepts of population structure and dynamics of many of the medically significant microorganisms have advanced significantly also. Currently, genetic identification of microbes is also within the reach of clinical microbiology laboratory professionals including those without specialized technology research interests. This review summarizes the possibilities for high-throughput molecular-microbiological typing in adequately equipped medical microbiology laboratories from both clinical and fundamental research perspectives. First, the development and application of methods for large-scale comparative typing of serially isolated microbial strains are discussed. The outcome of studies employing these methods allows for long-term epidemiologic surveillance of infectious diseases. Second, recent methods enable an almost nucleotide-by-nucleotide genetic comparison of smaller numbers of strains, thereby facilitating the identification of the genetic basis of, for instance, medically relevant microbiological traits. Whereas the first approach provides insights into the dynamic spread of infectious agents, the second provides insights into intragenomic dynamics and genetic functionality. The current state of technology is summarized, and future perspectives are sketched.
Pulsed-field gel electrophoresis typing was performed on a retrospective set of 129 Moraxella catarrhalis isolates obtained over a 20 month period from 70 children admitted to, or presenting at, the Erasmus University Medical Center, Rotterdam, The Netherlands. The mean age of the children (at the end of the study) was 2.5 years, with a range of 6 months to 15 years. Fifty-one different M. catarrhalis types were isolated from the hospitalized children, with 31 % (22/70) being infected with two particularly prevalent M. catarrhalis types. These two prevalent types also exhibited different protein profiles. The majority (72%; 16/22) of the children infected with these two predominant types had spent at least 1 week on two paediatric intensive care wards. No exacerbation of existing disease or new disease was observed in children who experienced M. catarrhalis type changes.
Nonchlamydial, nongonococcal urethritis (NCNGU) is suggested to be a sexually transmitted disease in men. NCNGU patients were compared to control subjects with regard to the presence of potentially infectious bacteria in the first void urine. Patients' pre- and post-antibiotic-treatment urine samples and two samples obtained 2 weeks apart from healthy volunteers, who did not receive antibiotic therapy, were analyzed with broad-spectrum PCR tests aiming at eubacterial small subunit rRNA genes. Restriction fragment length polymorphism analysis of the amplicons cloned from the mixtures of PCR products revealed that many different species of microorganisms were found to be colonizing the male urethra. We document here clear differences in the composition of the resident urethral flora between samples obtained from various individuals and between samples obtained at various points in time for a single individual. No major changes in population complexity were found upon antimicrobial treatment. In two of five patients a previously suggested pathogen (Mycoplasma genitalium or Haemophilus parainfluenzae) was accurately identified on the basis of DNA sequencing. No ubiquitous, azithromycin-sensitive organism was identified as a common pathogen in all patients, but up to 40% of all clones represented as-yet-unclassified bacterial species. Relatively often Pseudomonas spp. or Pseudomonas-like organisms were identified in the bacterial flora of patients. Interestingly, an as-yet-uncharacterized microbial species was identified as a negative predictor of NCNGU. This species was identified in all control subjects and was absent from all of the patient' samples (5 of 5 versus 0 of 5, P = 0.0079). This suggests that NCNGU might also be diagnosed by assessing the absence rather than the presence of certain bacterial species.
The objective of this study was to determine whether use of mupirocin nasal ointment for perioperative eradication of Staphylococcus aureus nasal carriage is effective in preventing the development of surgical site infections (SSIs). A randomized, double-blind, placebo-controlled design was used. Either mupirocin or placebo nasal ointment was applied twice daily to 614 assessable patients from the day of admission to the hospital until the day of surgery. A total of 315 and 299 patients were randomized to receive mupirocin and placebo, respectively. Eradication of nasal carriage was significantly more effective in the mupirocin group (eradication rate, 83.5% versus 27.8%). In the mupirocin group, the rate of endogenous S. aureus infections was 5 times lower than in the placebo group (0.3% and 1.7%, respectively; relative risk, 0.19; 95% confidence interval, 0.02-1.62). Mupirocin nasal ointment did not reduce the SSI rate (by S. aureus) or the duration of hospital stay.
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Staphylococcus aureus isolates (n = 225) from bovine teat skin, human skin, milking equipment, and bovine milk were fingerprinted by pulsed-field gel electrophoresis (PFGE). Strains were compared to assess the role of skin and milking equipment as sources of S. aureus mastitis. PFGE of SmaI-digested genomic DNA identified 24 main types and 17 subtypes among isolates from 43 herds and discriminated between isolates from bovine teat skin and milk. Earlier, phage typing (L. K. Fox, M. Gershmann, D. D. Hancock, and C. T. Hutton, Cornell Vet. 81:183-193, 1991) had failed to discriminate between isolates from skin and milk. Skin isolates from humans belonged to the same pulsotypes as skin isolates from cows. Milking equipment harbored strains from skin as well as strains from milk. We conclude that S. aureus strains from skin and from milk can both be transmitted via the milking machine, but that skin strains are not an important source of intramammary S. aureus infections in dairy cows. A subset of 142 isolates was characterized by binary typing with DNA probes developed for typing of human S. aureus. Typeability and overall concordance with epidemiological data were lower for binary typing than for PFGE while discriminatory powers were similar. Within several PFGE types, binary typing discriminated between main types and subtypes and between isolates from different herds or sources. Thus, binary typing is not suitable as replacement for PFGE but may be useful in combination with PFGE to refine strain differentiation.
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